title: Cycling probe technology to quantify and discriminate between wild-type varicella-zoster virus and Oka vaccine strains.
abstract: Rapid differentiation between wild-type varicella zoster virus (VZV) and Oka-vaccine (vOka) strains is important for monitoring side reactions of varicella vaccination. To develop a high-throughput molecular diagnostic method for the differentiation of wild-type VZV and vOka strains based on cycling probe technology. The primers were designed to amplify common sequences spanning a single nucleotide polymorphism (SNP) in gene 62 of VZV. DNA-RNA chimeric probes (cycling probes) were designed to detect the SNP at nucleotide 105705. The cycling probe real-time PCR assays for VZV wild-type and vOka strains specifically amplified plasmids containing target sequences that ranged between 10 and 1×10(6) copies per reaction. The inter- and intra-assay coefficients of variation were less than 5%. After initial validation studies, the clinical reliability of this method was evaluated using 38 swab samples that were collected from patients suspected of being zoster. Compared to the loop mediated isothermal amplification method, which is defined as the gold standard, cycling probe real-time PCR was highly sensitive and specific. The cycling probe real-time PCR technology is a reliable tool for differentiating between wild-type VZV and vOka strains in clinical samples.
authors.affiliation: Faculty of Clinical Engineering, Fujita Health University School of Health Sciences, Toyoake, Aichi, Japan. Electronic address: mihira@fujita-hu.ac.jp.
authors.forename: Masaru
authors.initials: M
authors.lastname: Ihira
authors.forename: Yuki
authors.initials: Y
authors.lastname: Higashimoto
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Kawamura
authors.forename: Ken
authors.initials: K
authors.lastname: Sugata
authors.forename: Masahiro
authors.initials: M
authors.lastname: Ohashi
authors.forename: Yoshizo
authors.initials: Y
authors.lastname: Asano
authors.forename: Tetsushi
authors.initials: T
authors.lastname: Yoshikawa
mesh_terms: D019433:Chickenpox Vaccine / Q000235:genetics*; D003937:Diagnosis, Differential; D006562:Herpes Zoster / Q000175:diagnosis* / Q000821:virology; D014645:Herpesvirus 3, Human / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry* / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods
chemical_list: D019433:Chickenpox Vaccine; D015345:Oligonucleotide Probes
keywords: Cycling probe; Molecular epidemiology; Oka vaccine; Real-time PCR; VZV